Determination of Dissolved Lignin by the Modified Pearl-Benson Nitrosation Method

1 Determination of Dissolved Lignin by the Modified Pearl-Benson (Nitrosation) Method 

10% Acetic Acid. Pipet 100 ml of reagent grade acetic acid into a 1000-ml volumetric flask and dilute to the mark with distilled water. 

2N Ammonium Hydroxide. Dilute 135 ml of concentrated ammonium hydroxide (28% as NH3) to 1000ml in a volumetric flask. This solution is stored in a Pyrex or borosilicate glass bottle and should be used only as long as it remains free from sediment when mixed with the sample of dissolved lignin in the procedure used to test reagent purity (Note 1). 

The water sample is allowed to stand for 12 to 24 h at room temperature (20-25 °C) after which the clear supernate is pipeted or decanted off. Samples which remain turbid after standing (i.e., those samples which show absorbance greater than 0.10 for a 10-cm path at 600 nm) should be filtered through a type AA Millipore filter (Millipore Filter Co., Bedford, MA). 

A 50-ml (Note 2) sample of water (settled or filtered through type AA Millipore filter) is placed in a 125-ml glass-stoppered Erlenmeyer flask and 1ml each of the acetic acid and sodium nitrite solutions are added from a buret or pipet. The solution is mixed by swirling and allowed to stand for 15 min at room temperature. Two ml of the ammonium hydroxide solution is added from a buret or pipet and mixed, and the solution is allowed to stand for 10 min at room temperature. The absorbance (Ax) is read at 430 nm in a cell with a 10-cm light path against a distilled water reference. 

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